1. Field of the Invention
The present invention provides methods for screening for voltage gated (VG)-selective inhibitors, novel VG-selective inhibitors, compositions containing the same, methods for inhibiting calcium entry into electrically non-excitable cells with said VG-selective inhibitors, methods for preventing proliferation of electrically non-excitable cells with said VG-selective inhibitors as well as methods of treating autoimmune diseases, preventing graft rejections, preventing apoptosis and treating cancer with the same.
2. Discussion of the Background
Cellular activation and proliferation are fundamental to myriad biologic processes. For example, appropriate activation of lymphocytes is the basis of immunity, while epithelial cell proliferation is required for wound healing. In other cases these same processes can be destructive. Inappropriate activation of lymphocytes produces autoimmune diseases while uncontrolled or dysregulated proliferation is the hallmark of cancer.
Recognition of the critical roles of cell activation and proliferation has spurred a quest to decipher the mechanisms regulating them in the hope of controlling them for therapeutic purposes. One of the earliest components of these regulatory mechanisms is an increase in the concentration of calcium within the cell. Virtually all of this increase is delivered to the cell interior, or cytosol, from the extracellular medium via calcium selective channels. To date there is no approach to the control of calcium entry in cells such as lymphocytes and epithelial cells that is applicable to the practice of medicine.
An increase in the cytosolic calcium concentration is so widespread and essential to cellular function that its regulation forms a dichotomous categorization of cell types. Cells in which calcium influx is regulated by electrical activity at the plasma membrane are called xe2x80x9celectrically excitablexe2x80x9d and are exemplified by neurons and muscle cells. The calcium channels in these cells are termed xe2x80x9cvoltage gatedxe2x80x9d (VG) because they are regulated primarily by the change in voltage across the plasma membrane. All other cells, including lymphocytes and epithelial cells, lack the type of electrical activity occurring in electrically excitable cells and so are named xe2x80x9celectrically non-excitablexe2x80x9d. The calcium channels in these types of cell are also referred to as VG channels by the inventors. To understand this invention, it is necessary to know that Ca2+ entry in these latter types of cells in conventionally believed to be conducted by non-voltage gated (NVG) channels.
Knowledge about voltage gated calcium channels in electrically excitable cells has been exploited profitably. Pharmacological modulation of these channels"" function is tremendously important in the practice of medicine; for example, calcium channel blockers are in widespread use in the treatment of epilepsy, hypertension, and angina pectoris. Unfortunately, such intervention is not yet available for calcium channels in electrically non-excitable cells. This deficiency likely reflects the fact that the mechanism by which calcium entry occurs has not been clearly identified.
Accordingly, one object of the present invention is to provide methods for screening for voltage gated (VG)-selective inhibitors.
A second object of the present invention is to provide novel VG-selective inhibitors.
A third object of the present invention is to provide novel compositions containing VG-selective inhibitors.
A fourth object of the present invention is to provide methods for inhibiting calcium entry into electrically non-excitable cells with said VG-selective inhibitors.
A fifth object of the present invention is to provide methods for preventing proliferation of electrically non-excitable cells, with said VG-selective inhibitors.
A sixth object of the present invention is to provide methods of treating autoimmune diseases, with said VG-selective inhibitors.
A seventh object of the present invention is to provide methods for preventing graft rejections, with said VG-selective inhibitors.
An eighth object of the present invention is to provide methods for preventing apoptosis.
A ninth object of the present invention is to provide methods for treating cancer.
The present inventors have now identified the current which mediates receptor-initiated increase in intracellular calcium in electrically non-excitable cells such as T cells: a novel Ca2+ voltage gated (VG) current and an electrically passive, non-voltage gated current. The inventors have discovered that identification of the proper current, and hence channel, in non-excitable cells is critical for the development of new pharmacologic agents that inhibit activation and proliferation of electrically non-excitable cells. The present inventors have now discovered that activation-associated entry of Ca2+ is mediated by the VG Ca2+ current, with properties similar to T type, low voltage activated Ca2+ currents, and that the non-voltage gated current is not required for activation-linked Ca2+ entry. Based on their discovery the above objects of the present invention were achieved.